Vascular Surgery Department, The First Affiliated Hospital of Xi'an Jiaotong University, 277 West Yanta Road, Xi'an, 710061, Shaanxi, China.
Nankai University School of Medicine, 94 Weijin Road, Tianjin, 300071, China.
Stem Cell Res Ther. 2021 Sep 9;12(1):497. doi: 10.1186/s13287-021-02561-w.
Adipose-derived mesenchymal stem cells (ADSC)-based therapy is an outstanding treatment strategy for ischaemic disease. However, the therapeutic efficacy of this strategy is not ideal due to the poor paracrine function and low survival rate of ADSCs in target regions. Platelet extracellular vesicles (PEVs) are nanoparticles derived from activated platelets that can participate in communication between cells. Accumulating evidence indicates that PEVs can regulate the biological functions of several cell lines. In the present study, we aimed to investigate whether PEVs can modulate the proangiogenic potential of ADSCs in vitro and in vivo.
PEVs were identified using scanning electron microscope (SEM), flow cytometry (FCM) and nanoparticle tracking analysis (NTA). The CCK8 assay was performed to detect proliferation of cells. Transwell and wound healing assays were performed to verify migration capacity of cells. AnnexinV-FITC/PI apoptosis kit and live/dead assay were performed to assess ADSCs apoptosis under Cocl-induced hypoxia condition. The underlying mechanisms by which PEVs affected ADSCs were explored using real time-PCR(RT-PCR) and Western blot. In addition, matrigel plug assays were conducted and mouse hindlimb ischaemic models were established to investigate the proangiogenic potential of PEV-treated ADSCs in vivo.
We demonstrated that ADSC could internalize PEVs, which lead to a series of biological reactions. In vitro, dose-dependent effects of PEVs on ADSC proliferation, migration and antiapoptotic capacity were observed. Western blotting results suggested that multiple proteins such as ERK, AKT, FAK, Src and PLCγ1 kinase may contribute to these changes. Furthermore, PEVs induced upregulation of several growth factors expression in ADSCs and amplified the proliferation, migration and tube formation of HUVECs induced by ADSC conditioned medium (CM). In in vivo experiments, compared with control ADSCs, the injection of PEV-treated ADSCs resulted in more vascularization in matrigel plugs, attenuated tissue degeneration and increased blood flow and capillary density in ischaemic hindlimb tissues.
Our data demonstrated that PEVs could enhance the proangiogenic potential of ADSCs in mouse hindlimb ischaemia. The major mechanisms of this effect included the promotion of ADSC proliferation, migration, anti-apoptosis ability and paracrine secretion.
脂肪间充质干细胞（ADSC）为基础的疗法是缺血性疾病的一种突出的治疗策略。然而，由于 ADSC 在靶区的旁分泌功能差和存活率低，这种策略的治疗效果并不理想。血小板细胞外囊泡（PEV）是来源于激活血小板的纳米颗粒，可以参与细胞间的通讯。越来越多的证据表明，PEV 可以调节几种细胞系的生物学功能。在本研究中，我们旨在研究 PEV 是否可以在体外和体内调节 ADSC 的促血管生成潜能。
使用扫描电子显微镜（SEM）、流式细胞术（FCM）和纳米颗粒跟踪分析（NTA）鉴定 PEV。CCK8 测定法用于检测细胞增殖。Transwell 和划痕愈合试验用于验证细胞迁移能力。AnnexinV-FITC/PI 凋亡试剂盒和活/死检测用于评估 Cocl 诱导的缺氧条件下 ADSC 的凋亡。使用实时 PCR（RT-PCR）和 Western blot 探索 PEV 影响 ADSC 的潜在机制。此外，进行了 Matrigel plugs 实验，并建立了小鼠后肢缺血模型，以研究体内 PEV 处理的 ADSC 的促血管生成潜能。
我们证明了 ADSC 可以内化 PEV，从而引发一系列生物学反应。在体外，观察到 PEV 对 ADSC 增殖、迁移和抗凋亡能力的剂量依赖性影响。Western blot 结果表明，多种蛋白，如 ERK、AKT、FAK、Src 和 PLCγ1 激酶，可能促成了这些变化。此外，PEV 诱导 ADSC 中多种生长因子表达的上调，并放大 ADSC 条件培养基（CM）诱导的 HUVEC 增殖、迁移和管形成。在体内实验中，与对照 ADSC 相比，注射 PEV 处理的 ADSC 导致 Matrigel plugs 中更多的血管生成，减轻组织退化，并增加缺血后肢组织中的血流和毛细血管密度。
我们的数据表明，PEV 可以增强小鼠后肢缺血模型中 ADSC 的促血管生成潜能。这种效应的主要机制包括促进 ADSC 的增殖、迁移、抗凋亡能力和旁分泌分泌。
